The following information is presented solely to assist the understanding of the reader, none of the information is admitted to describe prior art to the claims of the present invention.
This invention relates to compositions and methods for the introduction of a formulated nucleic acid into a cell for the expression of a peptide or polypeptide. It is useful for in vitro transfections and in vivo for gene therapy, for among other things administration of therapeutic proteins, polypeptides and peptides and for vaccination.
Non-viral administration of nucleic acid in vivo has been accomplished by a variety of methods. These include lipofectin/liposome fusion: Proc. Natl. Acad. Sci., Volume 84, pp. 7413-7417 (1993); polylysine condensation with and without adenovirus enhancement: Human Gene Therapy, Volume 3, pp. 147-154 (1992); and transferrin: transferrin receptor delivery of nucleic acid to cells: Proc. Natl. Acad. Sci., Volume 87, pp. 3410-3414 (1990). The use of a specific composition consisting of polyacrylic acid has been disclosed in WO 94/24983. Naked DNA has been administered as disclosed in WO 90/11092.
An important goal of gene therapy, as an initial step in the process of ultimately obtaining expression of a product encoded by a nucleic acid, is to effect the uptake of nucleic acid by cells. Uptake of nucleic acid by cells is dependent on a number of factors, one of which is the length of time during which a nucleic acid is in proximity to a cellular surface. For instance, after intramuscular (i.m.) administration of plasmid DNA in buffer, a marked reduction in gene expression is observed if the muscle is massaged, presumably due to DNA leakage out of the muscle either directly or via lymphatic vessels (Human Gene Therapy 4:151-159; 1993). Accordingly, it would be desirable to formulate nucleic acids with compounds which would retard the rate at which nucleic acids diffuse or are carried away from a site at which cellular uptake of the nucleic acid is desired. Further, these compounds would be suitable for administration to an organism by means such as injection while maintaining or regaining the physical characteristics necessary to increase cellular uptake of nucleic acids.
This invention features compositions and methods for enhancing the administration to and uptake of nucleic acids by an organism. An efficient strategy for enhancing nucleic acid delivery in vivo is to protect the nucleic acid from degradation, thereby maintaining the administered nucleic acid at the target site in order to further increase its cellular uptake. Also, for in vitro administration, increasing the effective concentration of the nucleic acid at the cell surface should increase the efficiency of transfection. The compositions of the present invention which are used to administer nucleic acid comprise a compound which protects the nucleic acid and/or prolongs the localized bioavailability of the nucleic acid when administered to an organism in vivo, or in vitro in cell culture.
In connection with the compounds and compositions of this invention, the term xe2x80x9cprotectsxe2x80x9d or xe2x80x9cprotectivexe2x80x9d refers to an effect of the interaction between such a compound and a nucleic acid such that the rate of degradation of the nucleic acid is decreased in a particular environment. Such degradation may be due to a variety of different factors, which specifically include the enzymatic action of a nuclease. The protective action may be provided in different ways, for example, by exclusion of the nuclease molecules or by exclusion of water.
By xe2x80x9cprolong the localized bioavailability of a nucleic acidxe2x80x9d is meant that a nucleic acid when administered to an organism in a composition comprising such a compound will be available for uptake by cells for a longer period of time than if administered in a composition without such a compound, for example when administered in a formulation such as a saline solution. This increased availability of nucleic acid to cells could occur, for example, due to increased duration of contact between the composition containing the nucleic acid and a cell or due to protection of the nucleic acid from attack by nucleases. The compounds which prolong the localized bioavailability of a nucleic acid are suitable for internal administration.
By xe2x80x9csuitable for internal administrationxe2x80x9d is meant that the compounds are suitable to be administered within the tissue of an organism, for example within a muscle or within a joint space, intradermally or subcutaneously. Other forms of administration which may be utilized are topical, oral, pulmonary, nasal and mucosal; for example, buccal, vaginal or rectal. Properties making a compound suitable for internal administration can include, for example, the absence of a high level of toxicity to the organism as a whole.
By xe2x80x9cnucleic acidxe2x80x9d is meant both RNA and DNA including: cDNA, genomic DNA, plasmid DNA or condensed nucleic acid, nucleic acid formulated with cationic lipids, nucleic acid formulated with peptides, cationic polymers, RNA or mRNA. In a preferred embodiment, the nucleic acid administered is plasmid DNA which comprises a xe2x80x9cvectorxe2x80x9d.
A xe2x80x9cvectorxe2x80x9d is a nucleic acid molecule incorporating sequences encoding therapeutic product(s) as well as, various regulatory elements for transcription, translation, transcript stability, replication, and other functions as are known in the art.
A xe2x80x9ctranscript stabilizerxe2x80x9d is a sequence within the vector which contributes to prolonging the half life (slowing the elimination) of a transcript.
xe2x80x9cPost-translational processingxe2x80x9d means modifications made to the expressed gene product. These may include addition of side chains such as carbohydrates, lipids, inorganic or organic compounds, the cleavage of targeting signals or propeptide elements, as well as the positioning of the gene product in a particular compartment of the cell such as the mitochondria, nucleus, or membranes. The vector may comprise one or more genes in a linear or circularized configuration. The vector may also comprise a plasmid backbone or other elements involved in the production, manufacture, or analysis of a gene product.
An xe2x80x9cexpression vectorxe2x80x9d is a vector which allows for production of a product encoded for by a nucleic acid sequence contained in the vector. For example, expression of a particular growth factor protein encoded by a particular gene.
A xe2x80x9cDNA vectorxe2x80x9d is a vector whose native form is a DNA molecule. A xe2x80x9cviral vectorxe2x80x9d is a vector whose native form is as the genomic material of a viral particle.
A xe2x80x9cgene productxe2x80x9d means products encoded by the vector. Examples of gene products include mRNA templates for translation, ribozymes, antisense RNA, proteins, glycoproteins, lipoproteins and phosphoproteins. The nucleic acid may be associated with a targeting ligand to effect targeted delivery.
A xe2x80x9ctargeting ligandxe2x80x9d is a component of the carrier or vehicle or a moiety (a molecule or group) attached to a protective, interactive, non-condensing compound which binds to receptors, with an affinity for the ligand, on the surface or within compartments of a cell for the purpose of enhancing uptake or intracellular trafficking of the vector. Galactosyl residues, e.g., Tris-galactosyl residues, carnitine derivatives, mannosyl residues, e.g., mannose-6-phosphate, monoclonal and polyclonal antibodies, peptide ligands, and DNA-binding proteins represent non-limiting examples of targeting ligands which can be used to enhance uptake.
xe2x80x9cTargeted deliveryxe2x80x9d involves the use of targeting ligands which specifically enhance translocation of a nucleic acid to specific tissues or cells. Examples of cells which may be targeted include, but are not limited to, antigen-presenting cells, hepatocytes, myocytes, eptithelial cells, endothelial cells, and cancer cells.
A xe2x80x9ctargetxe2x80x9d is a specific organ, tissue, cell, or cellular region for which uptake of a vector and expression of a gene product is intended.
xe2x80x9cUptakexe2x80x9d means the translocation of the vector from the extracellular to intracellular compartments. This can involve receptor mediated processes, fusion with cell membranes, endocytosis, potocytosis, pinocytosis or other translocation mechanisms. The vector may be taken up by itself or as part of a complex.
xe2x80x9cBindingxe2x80x9d is an intermediate step in uptake of some complexes involving a high-affinity interaction between a targeting ligand and a surface receptor on a target cell.
xe2x80x9cIntracellular traffickingxe2x80x9d is the translocation of the vector within the cell from the point of uptake to the nucleus where expression of a gene product takes place. Alternatively, cytoplasmic expression of a nucleic acid construct utilizing, for example, a T7 polymerase system may be accomplished. Various steps in intracellular trafficking include endosomal release and compartmentalization of the vector within various extranuclear compartments, and nuclear entry.
xe2x80x9cEndosomal releasexe2x80x9d is the egress of the vector from the endosome after endocytosis. This is an essential and potentially rate limiting step in the trafficking of vectors to the nucleus. A lytic peptide may be used to assist in this process.
A xe2x80x9clytic peptidexe2x80x9d is a peptide which functions alone or in conjunction with another compound to penetrate the membrane of a cellular compartment, particularly a lysosomal or endosomal compartment, to allow the escape of the contents of that compartment to another cellular compartment such as the cytosolic and/or nuclear compartment.
xe2x80x9cCompartmentalizationxe2x80x9d is the partitioning of vectors in different compartments within a defined extracellular or intracellular space. Significant extracellular compartments may include, for example, the vascular space, hair follicles, interstitial fluid, synovial fluid, cerebral spinal fluid, thyroid follicular fluid. Significant intracellular compartments may include endosome, potosome, lysosome, secondary lysosome, cytoplasmic granule, mitochondria, and the nucleus.
xe2x80x9cNuclear entryxe2x80x9d is the translocation of the vector across the nuclear membrane into the nucleus where the gene may be transcribed.
xe2x80x9cEliminationxe2x80x9d is the removal or clearance of materials (vectors, transcripts, gene products) from a specific compartment over time. This term may be used to reflect elimination from the body, the vascular compartment, extracellular compartments, or intracellular compartments. Elimination includes translocation (excretion) from a particular compartment or biotransformation (degradation).
The compounds which protect the nucleic acid and/or prolong the localized bioavailability of a nucleic acid may achieve one or more of the following effects, due to their physical, chemical or Theological properties: (1) Protect nucleic acid, for example plasmid DNA, from nucleases due to steric, viscosity, or other effects; (2) increase the area of contact between nucleic acid, such as plasmid DNA, through extracellular matrices and over cellular membranes, into which the nucleic acid is to be taken up; (3) concentrate nucleic acid, such as plasmid DNA, at cell surfaces due to water exclusion; (4) indirectly facilitate uptake of nucleic acid, such as plasmid DNA, by disrupting cellular membranes due to osmotic, hydrophobic or lytic effects; and (5) indirectly facilitate uptake of nucleic acids by allowing diffusion of protected nucleic acid chains through tissue at the administration site.
The following polymers, oils and surfactants may be suitable for use as compounds which prolong the localized bioavailability of a nucleic acid: polyvinylpyrrolidones; polyvinylalcohols; propylene glycols; polyethylene glycols; polyvinylacetates; poloxamers (Pluronics)(block copolymers of propylene oxide and ethylene oxide, relative amounts of the two subunits may vary in different poloxamers); poloxamines (Tetronics); ethylene vinyl acetates; celluloses, including salts of carboxymethylcelluloses, methylcelluloses, hydroxypropylcelluloses, hydroxypropylmethylcelluloses; salts of hyaluronates; salts of alginates; heteropolysaccharides (pectins); dextrans; chitosans; phosphatidylcholines (lecithins); miglyols; polylactic acid; polyhydroxybutyric acid. As indicated below, certain of these compounds may be used as protective, interactive, non-condensing compounds and others as sustained release compounds, while some may be used in either manner under the respectively appropriate conditions.
These substances may be prepared as solutions, suspensions, gels, emulsions or microemulsions of a water/oil (w/o), water/oil/water (w/o/w), oil/water (o/w) or oil/water/oil (o/w/o) type. Oil suspensions of lyophilized nucleic acid, such as plasmid DNA may be utilized. Carriers for these oil suspensions include, but are not limited to, sesame oil, cottonseed oil, soybean oil, lecithins, Tweens, Spans and Miglyols.
By xe2x80x9csolutionsxe2x80x9d is meant water soluble polymers and/or surfactants in solution with nucleic acids.
By xe2x80x9csuspensionsxe2x80x9d is meant water insoluble oils containing suspended nucleic acids.
By xe2x80x9cgelsxe2x80x9d is meant high viscosity polymers containing nucleic acids.
By xe2x80x9cemulsionxe2x80x9d is meant a dispersed system containing at least two immiscible liquid phases. Emulsions usually have dispersed particles in the 0.1 to 100 micron range. They are typically opaque and thermodynamically unstable. Nucleic acids in the water phase can be dispersed in oil to make a w/o emulsion. This w/o emulsion can be dispersed in a separate aqueous phase to yield a w/o/w emulsion. Alternatively, a suitable oil could be dispersed in an aqueous phase to form an o/w emulsion. A xe2x80x9cmicroemulsionxe2x80x9d has properties intermediate to micelles and emulsions and is characterized in that they are homogenous, transparent and thermodynamically stable. They form spontaneously when oil, water, surfactant and cosurfactant are mixed together. Typically, the diameter of the dispersed phase is 0.01 to 0.1 microns, usually of the w/o and o/w type.
Some compounds which prolong the bioavailability of a nucleic acid may also interact or associate with the nucleic acid by intermolecular forces and/or valence bonds such as: Van der Waals forces, ion-dipole interactions, ion-induced dipole interactions, hydrogen bonds, or ionic bonds. These interactions may serve the following functions: (1) Stereoselectively protect nucleic acids from nucleases by shielding; (2) facilitate the cellular uptake of nucleic acid by xe2x80x9cpiggyback endocytosisxe2x80x9d. Piggyback endocytosis is the cellular uptake of a drug or other molecule complexed to a carrier that may be taken up by endocytosis. CV Uglea and C Dumitriu-Medvichi, Medical Applications of Synthetic Oligomers, In: Polymeric Biomaterials, Severian Dumitriu ed., Marcel Dekker, Inc., 1993, incorporated herein by reference.
To achieve the desired effects set forth it is desirable, but not necessary, that the compounds which prolong the bioavailability of a nucleic acid have amphiphilic properties; that is, have both hydrophilic and hydrophobic regions. The hydrophilic region of the compounds may associate with the largely ionic and hydrophilic regions of the nucleic acid, while the hydrophobic region of the compounds may act to retard diffusion of nucleic acid and to protect nucleic acid from nucleases.
Additionally, the hydrophobic region may specifically interact with cell membranes, possibly facilitating endocytosis of the compound and thereby also of nucleic acid associated with the compound. This process may increase the pericellular concentration of nucleic acid.
Agents which may have amphiphilic properties and are generally regarded as being pharmaceutically acceptable are the following: polyvinylpyrrolidones; polyvinylalcohols; polyvinylacetates; propylene glycol; polyethylene glycols; poloxamers (Pluronics); poloxamines (Tetronics); ethylene vinyl acetates; methylcelluloses, hydroxypropylcelluloses, hydroxypropylmethylcelluloses; heteropolysaccharides (pectins); chitosans; phosphatidylcholines (lecithins); miglyols; polylactic acid; polyhydroxybutyric acid; xanthan gum. Also, copolymer systems such as polyethylene glycol-polylactic acid (PEG-PLA), polyethylene glycol-polyhydroxybutyric acid (PEG-PHB), polyvinylpyrrolidone-polyvinylalcohol (PVP-PVA), and derivatized copolymers such as copolymers of N-vinyl purine (or pyrimidine) derivatives and N-vinylpyrrolidone. However, not all of the above compounds are protective, interactive, non-condensing compounds as described below.
In a first aspect, the invention provides compositions for the delivery of a nucleic acid to a cell. Such a composition includes a protective, interactive, non-condensing, amphiphilic compound (PINC) and a nucleic acid molecule. The PINC enhances the delivery of the nucleic acid molecule to mammalian cells in vivo, and preferably the nucleic acid molecule includes a coding sequence for a gene product to be expressed in the cell. In many cases, the relevant gene product is a polypeptide or protein.
In a related aspect, the invention also provides compositions for delivery of a nucleic acid molecule to a cell. As in the preceding aspect, the composition includes a PINC and a nucleic acid molecule. Also as in the preceding aspect, preferably the PINC is used under conditions so that the PINC does not form a gel, or so that no gel form is present at the time of administration at about 30-40xc2x0 C. Thus, in these compositions, the PINC is present at a concentration of 30% (w/v) or less. In certain preferred embodiments, the PINC concentration is still less, for example, 20% or less, 10% or less, 5% or less, or 1% or less. Thus, these compositions differ in compound concentration and functional effect from uses of these or similar compounds in which the compounds are used at higher concentrations, for example in the ethylene glycol mediated transfection of plant protoplasts, or the formation of gels for drug or nucleic acid delivery. In general, the PINCs are not in gel form in the conditions in which they are used as PINCs, though certain of the compounds may form gels under some conditions.
In connection with the protective, interactive, non-condensing compounds for these compositions, the term xe2x80x9cnon-condensingxe2x80x9d means that an associated nucleic acid is not condensed or collapsed by the interaction with the PINC at the concentrations used in the compositions. Thus, the PINCs differ in type and/or use concentration from such condensing polymers. Examples of commonly used condensing polymers include polylysine, and cascade polymers (spherical polycations).
Also in connection with such compounds and an associated nucleic acid molecule, the term xe2x80x9cenhances the deliveryxe2x80x9d means that at least in conditions such that the amounts of PINC and nucleic acid is optimized, a greater biological effect is obtained than with the delivery of nucleic acid in saline. Thus, in cases where the expression of a gene product encoded by the nucleic acid is desired, the level of expression obtained with the PINC:nucleic acid composition is greater than the expression obtained with the same quantity of nucleic acid in saline for delivery by a method appropriate for the particular PINC/coding sequence combination.
In preferred embodiments of the above compositions, the gene product is a ribonucleic acid molecule, a polypeptide, or protein.
Also in preferred embodiments of the above compositions, the PINC is polyvinyl pyrrolidone (PVP), polyvinyl alcohol (PVA), a PVP-PVA co-polymer, N-methyl-2-pyrrolidone (NM2P), ethylene glycol, or propylene glycol. In compositions in which a Poloxamer (Pluronics) is used, the nucleic acid is preferably not a viral vector, i.e., the nucleic acid is a non-viral vector.
In other preferred embodiments, the PINC is bound with a targeting ligand. Such targeting ligands can be of a variety of different types, including but not limited to galactosyl, residues, fucosal residues, mannosyl residues, carntitine derivatives, monoclonal antibodies, polyclonal antibodies, peptide ligands, and DNA-binding proteins. The targeting ligands may bind with receptors on cells such as antigen-presenting cells, hepatocytes, myocytes, epithelial cells, endothelial cells, and cancer cells.
In connection with the association of a targeting ligand and a PINC, the term xe2x80x9cbound withxe2x80x9d means that the parts have an interaction with each other such that the physical association is thermodynamically favored, representing at least a local minimum in the free energy function for that association. Such interaction may involve covalent binding, or non-covalent interactions such as ionic, hydrogen bonding, van der Waals interactions, hydrophobic interactions, and combinations of such interactions.
While the targeting ligand may be of various types, in one embodiment the ligand is an antibody. Both monoclonal antibodies and polyclonal antibodies may be utilized.
The nucleic acid may also be present in various forms. Preferably the nucleic acid is not associated with a compound(s) which alter the physical form, however, in other embodiments the nucleic acid is condensed (such as with a condensing polymer), formulated with cationic lipids, formulated with peptides, or formulated with cationic polymers.
As the compositions are useful for delivery of a nucleic acid molecule to cells in vivo, in a related aspect the invention provides a composition at an in vivo site of administration. In particular this includes at an in vivo site in a mammal.
In preferred embodiments the nucleic acid molecule includes a sequence encoding a gene product. Also in preferred embodiments, the site of administration is in an interstitial space or a tissue of an animal, particularly of a mammal.
The invention also provides methods for using the above compositions. Therefore, in further related aspects, methods of administering the compositions are provided in which the composition is introduced into a mammal, preferably into a tissue or an interstitial space.
Various methods of delivery may be utilized, such as are known in the art, but in preferred embodiments, the composition is introduced into the tissue or interstitial space by injection. The compositions may also be delivered to a variety of different tissues, but in preferred embodiments the tissue is muscle or tumor.
In another related aspect, the invention provides methods for treating a mammalian condition or disease by administering a therapeutically effective amount of a composition as described above. In preferred embodiments, the disease or condition is a cancer.
A xe2x80x9ctherapeutically effective amountxe2x80x9d of a composition is an amount which is sufficient to cause at least temporary relief or improvement in a symptom or indication of a disease or condition. Thus, the amount is also sufficient to cause a pharmacological effect. The amount of the composition need not cause permanent improvement or improvement of all symptoms or indications.
In another aspect of the invention, the compound which prolongs the bioavailability of a nucleic acid is a sustained-release compound which may be administered to an organism or to cells in culture. The sustained-release compound containing a nucleic acid is administered to the tissue of an organism, for example, by injection. In one embodiment the tissue is preferably muscle tissue. In another embodiment the tissue is preferably a joint space. In another embodiment the tissue is preferably a tumor.
By xe2x80x9csustained-release compoundxe2x80x9d is meant a substance with a viscosity above that of an isotonic saline solution (150 mM NaCl) containing a nucleic acid; for example, DNA in saline at 1 mg/ml has a viscosity of 3.01 mPaxc2x7sec, DNA in saline at 2 mg/ml has a viscosity of 3.26 mPaxc2x7sec, DNA in saline at 3 mg/ml has a viscosity of 5.85 mPaxc2x7sec (Viscosity measurements were performed at 25xc2x0 C. in a Brookfield DV-III Rheometer with a No. 40 Spindle at 75 rpm for 30 minutes).
Preferably the sustained-release compound has a viscosity in the range of about 0.1-20,000 mPaxc2x7sec above that of a formulation in which isotonic saline is the carrier for a nucleic acid. More preferably the range is about 0.1-5000 mPaxc2x7sec above that of a formulation in which isotonic saline is the carrier for a nucleic acid. Even more preferably the range is about 0.1-1000 mPaxc2x7sec above that of a formulation in which isotonic saline is the carrier for a nucleic acid.
By xe2x80x9csustained-releasexe2x80x9d is meant that nucleic acid is made available for uptake by surrounding tissue or cells in culture for a period of time longer than would be achieved by administration of the nucleic acid in a less viscous medium, for example, a saline solution.
In another embodiment, the compound which prolongs the bioavailability of a nucleic acid is a thermo-reversible gel.
By xe2x80x9cthermo-reversible gelxe2x80x9d is meant a gel which is substantially liquid at temperatures below about 30xc2x0 C. but forms a gel at temperatures above about 30xc2x0 C. Administration of the thermo-reversible gel by, for example, injection is thereby facilitated if the gel is cooled so that it is in a substantially liquid state when injected. However, upon contact with the tissue of an organism which is at a temperature of above about 30xc2x0 C. the viscosity of the thermo-reversible gel increases, thereby increasing the localized bioavailability of a nucleic acid formulated with the thermo-reversible gel.
In another embodiment of the present invention, the molecules of the compound which prolongs the localized bioavailability of a nucleic acid tend to orient themselves in the direction of an induced flow and as an applied force causing the flow is increased and the resistance of the compound to flow is decreased, lowering an initial viscosity of the compound. When the applied force is removed, the compound substantially reverts to its initial viscosity. In a preferred embodiment the compound utilized is a salt of carboxymethylcellulose, such as sodium carboxymethylcellulose. Sodium carboxymethylcellulose has been used by the cosmetics, food, and pharmaceutical industries as a stabilizer, thickener, gelling agent, suspending agent, and a lubricant. Sodium carboxymethyl cellulose is an approved pharmaceutical excipient.
In another embodiment, the compound which prolongs the bioavailability of a nucleic acid is polyvinylpyrrolidone (PVP). PVP is a polyamide that forms complexes with a wide variety of substances and is chemically and physiologically inert. Specific examples of suitable PVP""s are Plasdone-C(copyright)15, MW 10,000 and Plasdone-C(copyright)30, MW 50,000.
In another embodiment the compound which prolongs the bioavailability of a nucleic acid is an oily suspension. By xe2x80x9coily suspensionxe2x80x9d is meant a coarse dispersion containing finely divided insoluble material suspended in a liquid medium. These formulations include: nucleic acids, polymers, peptides or sugars and are dispersed with the aid of a dispersing agent, such as a surfactant in a suitable vehicle such as an oil. For example, DNA/PVP powder blend in Miglyol with 0.1% Tween-80, DNA/PVP powder blend in sesame oil with 0.1% Tween-80, DNA/lactose powder blend in Miglyol with 0.1% Tween-80, DNA complex powder blends in Miglyol with 0.1% Tween-80, where the DNA complex could comprise condensed DNA complexes such as DNA:polymer or DNA:peptide.
In another embodiment the compound which prolongs the bioavailability of a nucleic acid is a water-in-oil microemulsion. Examples would include: lecithin:sesame oil:butanol (surfactant/oil/cosurfactant) as the oil phase with DNA in saline as the water phase; lecithin:sesame oil:butanol (surfactant/oil/cosurfactant) as the oil phase with DNA complex saline as the water phase.
In another embodiment the compound which prolongs the bioavailability of a nucleic acid is a hydrogel. Nucleic acids may be loaded into hydrogels by placing swellable hydrogel systems in nucleic acid solutions. Swellable hydrogels include but are not limited to hydroxyethylmethacrylate (HEMA), polyethyleneglycolmethacrylate (PEGMA), cellulose ether hydrogels, comprising cross-linked hydroxypropyl cellulose, methyl cellulose, and hydroxypropylmethyl cellulose; calcium-crosslinked alginate; crosslinked polyvinyl alcohols and Poloxamers (Pluronics).
In another embodiment the compound which prolongs the bioavailability of a nucleic acid is a cationic polymer, such as Eudragit, Chitosan and Poloxamines (Tetronics).
In another embodiment the compound which prolongs the bioavailability of a nucleic acid is a surfactant which forms micelles, such as Tween 80.
In another embodiment the uptake of nucleic acids in vitro, for example, cells in tissue culture is enhanced by the use of the compounds disclosed herein.
Other and further objects, features, and advantages will be apparent from the following description of the presently preferred embodiments in the invention.